Synthetic sgRNA and crRNA Service

Synthetic CRISPR RNA and Cas9 Nuclease Reagents for Gene Editing

CRISPR/Cas9-mediated gene editing is a powerful technique that allows you to create knock-in/out mutations in any gene and any cell. Using the ribonucleoprotein (RNP) system has many advantages over other forms of gene editing.

Advantages

DNA free

Detectable at high levels shortly after transfection

Quickly cleared from the cell for less off-target effects

Highly efficient even in hard-to-transfect cells

Best for in vivo studies, studies show that sgRNA has better stability than crRNA:tracrRNA when duplexed with Cas9

GenScript provides both unmodified sgRNA and modified sgRNAs. Modified sgRNAs with 2'-O-methyl and phosphorothioate modifications at the first three 5' and 3' terminal residues are recommended for better stability and editing efficiency.

*Deliver in as short as 8 business days.

Product

Length

Quantity

Pricing

GenCRISPR sgRNA

97 to 103 nt

4 nmol

$259

Modified GenCRISPR sgRNA (2'-O-methyl and phosphorothioate modifications at the first three 5' and 3' terminal RNA residues)

97 to 103 nt

4 nmol

$359

CRISPR/Cas9 crRNAs

GenScript CRISPR RNAs (crRNAs) are short 36 nt RNA oligonucleotides which contain a short 20 nt sequence which guide the CRISPR/Cas9 complex to genomic targets for gene editing.

*Deliver in as short as 8 business days.

Product

Length

Quantity

Pricing

GenCRISPR crRNA

20 nt

2 nmol

$95

GenCRISPR crRNA

20 nt

10 nmol

$125

GenCRISPR crRNA

20 nt

20 nmol

$225

CRISPR/Cas9 tracrRNAs

GenScript CRISPR/Cas9 trans-activating crRNAs (tracrRNAs) are 67 nt RNA oligonucleotides which together with the crRNA and Cas9 nuclease, form the activated CRISPR/Cas9 ribonucleoprotein complex.

*In-stock

Product

Quantity

Pricing

GenCRISPR tracrRNAs

5 nmol

$95

GenCRISPR tracrRNAs

10 nmol

$125

GenCRISPR tracrRNAs

20 nmol

$225

Nuclease-Free Annealing Buffer

For annealing tracrRNA and crRNA.

Product

Concentration

Quantity

Pricing

Nuclease-Free Annealing Buffer

5X

100 µL

$5

Nuclease-Free Annealing Buffer

5X

1 mL

$10

HPRT Primer Mix

HPRT positive control is highly recommended for testing editing efficiency.

CRISPR/Cas9 Control Kit

GenScript offers pre-duplexed human HPRT crRNA:tracrRNA as positive controls for your CRISPR experiments. HPRT (hypoxanthine phosphoribosyltransferase) is a housekeeping gene and commonly used control has already been pre-validated for efficient CRISPR cleavage. The kit comes with an HPRT primer mix for PCR analysis.

*In-stock

Product

Pricing

GenCRISPR/Cas9 Control Kit

$195

Synthetic crRNA/tracrRNA

CRISPR/Cas9 ribonucleoprotein (RNP) complexes are composed of a CRISPR RNA (crRNA), trans-activating crRNA (tracrRNA) duplex and Cas9 protein. The crRNA consists of a 20 nt sequence that is complementary to a genomic target. crRNA sequence design utilizes standard gRNA design principles and sequences can be easily selected from the GenScript gRNA database or customized using the GenScript gRNA design tool. When complexed with tracrRNA and Cas9 protein, a double strand break will be created at a locus complementary to the 20 nt guide RNA sequence, 3-4 bp upstream from a protospacer adjacent motif (PAM) sequence (5'-NGG-3'). Unlike traditional CRISPR Plasmids, CRISPR/Cas9 ribonucleoproteins are delivered as intact complexes, and do not require cellular expression.

CRISPR RNA/Cas9 Protein Workflow

Only three steps are required for using synthetic CRISPR RNA oligos:

Incubate the crRNA and tracrRNA with the annealing buffer

Incubate the duplexed crRNA:tracrRNA with Cas9 protein to form the active CRISPR/Cas9 ribonucleoprotein. Optionally, an HDR (homology dependent repair) template can also be co-delivered with the Cas9 RNP for targeted knock-in.

Deliver the ribonucleoprotein mix into the cells

Prior to experimentation, we recommend first testing your CRISPR/Cas9 complex in vitro and in test cell lines to ensure cleavage and transfection efficiency. Delivery of the CRISPR/Cas9 complex into cells is typically performed by electroporation or via lipid-mediated transfection.

All-in-one vector systems have two main advantages:

Cells only need to be transfected once.

gRNA/Cas9 expression is driven in an ideal 1:1 ratio.

Dual vectors, where Cas9 and gRNA are expressed independently on separate constructs, are more suitable if you plan to express multiple gRNAs for multiplex targeting. For these applications, Cas9 should first be stably expressed in the cell line, after which the cells can be transfected with different gRNA vectors to generate a cell pool.

Keep the crRNA and tracrRNA oligonucleotides tightly sealed at -20℃ prior to use and avoid repeated freeze-thaw cycles. We recommend working in a sterile environment, using RNase-free pipette tips and tubes.

For 5X Annealing buffer:

1. Centrifuge tubes before opening to ensure RNA oligos are at the bottom of the tube.

2. Resuspend both oligos in Nuclease-Free water to reach the appropriate final concentration, for example,100µM.